W0173

Structure Determination of Hydroxyethyl Thiazole Kinase and the Importance of Low Resolution Data. Nino Campobasso¹, Ulricke Breitinger¹, Yi Zhang², Tadhg P. Begley², and Steve Ealick¹, ¹Section of Biochemistry, Molecular, and Cellular Biology & ²Department of Chemistry, Cornell University, Ithaca NY, 14850

Hydroxyethyl thiazole kinase is a salvage enzyme in the biosynthesis of thiamin (Vitamin B1). The kinase structure was determined to 1.6Å resolution by single isomorphous replacement anomalous scattering. We will present the structure of hydroxyethyl thiazole kinase and show the importance of low resolution data for solvent flattening.

Hydroxyethyl thiazole kinase has 272 amino acids and forms a trimer in solution. The enzyme crystallizes in the spacegroup R3 with unit cell parameters of 77.5Å x 77.5Å x 230.4Å with two monomers in the asymmetric unit. On a frozen crystal, native 1.6Å resolution X-ray diffraction data were collected at CHESS, beamline A1, using a 2k x 2k CCD area detector. A crystal of Hydroxyethyl thiazole kinase was derivatized with a mercurial compound, and x-ray diffraction data to 2.2Å resolution were also collected at CHESS. Both data sets were 90% complete and produced interpretable difference Patterson maps and good phasing statistics; however, the electron density was not interpretable. Closer observation of the x-ray data showed that the data were only 80% complete from infinity - 8Å resolution. Upon collecting a complete low resolution data set, the electron density was interpretable.